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Plant regeneration from CSC of E. alba. a Cells collected from CSC (Bar 10 mm); b embryogenic callus (Bar 10 mm); c plantlets obtained from MS medium supplemented with ABA (0.5 mg L⁻¹) in association with BA (0.75 mg L⁻¹) (Bar 10 mm); d root induction on 1/2MS medium plus 1.0 mg L⁻¹ IAA (Bar 20 mm); (e) Regenerated plantlets acclimatized in garden soil, sand and vermicompost (1:1:1; v/v) (Bar 30 mm)
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The present research focused on enhancing the production of wedelolactone through cell suspension culture (CSC) in Eclipta alba (L.) Hassk. With an aim of attaining a sustainable CSC, various plant growth regulators, elicitors and agitation speed were examined. Nodal segments of in vitro propagated plantlets induced the maximum percentage (93.47 ±...
Citations
... Previous research has suggested that treating elicitors at the end of the log/exposure or stationary phase of cell growth increases the production of bioactive molecules in Hypericum perforatum (Wang et al. 2015), Ophiorrhiza mungos (Deepthi and Satheeshkumar 2016), Eclipta alba (Salma et al. 2018), Pueraria candollei var. mirifica (Udomsin et al. 2020), Gymnema sylvestre (Mahendran et al. 2021a, b), and Galega officinalis (Khezri et al. 2022). ...
Enicostema axillare (Poir. ex Lam.) A. Raynal is an immense medicinal plant that has been extensively used to treat malaria, diabetes, stomachache, and fever in traditional medicine. This plant contains high amounts of secoiridoid glycosides (gentiopicroside and swertiamarin), which have numerous health benefits. However, no leaf callus or cell suspension culture has been established for E. axillare for the production of beneficial metabolites. As a result, the goal of this work was to standardize an efficient and robust method for callus induction, callus and cell biomass, as well as swertiamarin and gentiopicroside accumulation. The highest callus induction frequency (85.40 ± 0.51%) and formation of leaf into callus with a mean callus biomass (1.283 ± 0.02 g/explant fresh weight (FW) and 0.176 ± 0.00 g/explant dry weight (DW)) and metabolite contents (swertiamarin 0.53 ± 0.01 mg/g and gentiopicroside 0.64 ± 0.05 mg/g) were obtained on Murashige and Skoog (MS) medium augmented with 2,4-D (1.0 mg/L) + Kin (0.5 mg/L) after 12 weeks of culture. The induced leaf callus was studied for cell suspension growth kinetics (biomass) and metabolite production. After 60th days of culture, the maximum of the leaf cell suspension (5.25 ± 0.05 g FW (50 mL medium) and 0.39 ± 0.00 g DW (50 mL medium) and secoiridoid glycosides (gentiopicroside, 1.44 ± 0.05 mg/g, and swertiamarin, 0.88 ± 0.01 mg/g DW) were obtained, which corresponded to 4.19 and 36 times greater than the lag phase (after 10th days). Cultured leaf cell suspensions were treated with methyl jasmonate (MJ) and salicylic acid (SA) at 50, 100 and 200 μM after 60th day of cell growth and evaluated for swertiamarin and gentiopicroside contents after 24 h of treatment. The greatest swertiamarin (2.43 ± 0.05 mg/g DW, 2.76-fold) and gentiopicroside (2.41 ± 0.09 mg/g DW, 1.67-fold) levels were observed in the treatments with 200 and 100 μM SA elicitor, respectively. These findings can lead to the use of this plant as an alternative source to improve the commercial production of secoiridoid glycosides in large-scale cultures.
... Earlier studies demonstrated that elicitors are very effective for the enhancement of cell growth and increase metabolites accumulation. Furthermore, the type of elicitors, concentration and exposure period favours the biomass accumulation in cell suspension culture (Salma et al. 2018;Zare-Hassani et al. 2019;Açıkgöz, 2020). ...
Gymnemic acids (a group of triterpenoid saponins) found in Gymnema sylvestre (Retz.) R.Br. ex Sm. works as the main hypoglycaemic active components. These can be the potential active pharmaceutical ingredient (API) to be used by pharmaceutical industries in modern medicines against diabetes. The present study aims to investigate the effectiveness of sodium nitroprusside (SNP) treatment for enhancement of cell suspension culture biomass and to quantify the production of deacylgymnemic acid, gymnemagenin, gymnemic acid IV and gymnemic acid XVII contents. Callus was obtained from in vitro derived leaves of G. sylvestre on MS medium fortified with 3.0 mg/L 2, 4-d (2, 4-dichlorophenoxyacetic acid) and 2.0 mg/L Kn (Kinetin), and the same were used further to produce cell suspension cultures. Cell suspensions were exposed to different concentrations of SNP (5, 10, 20 and 40 μM) and data were collected at 20, 30 and 40 days. Out of the tested concentrations, 20 μM SNP had the highest level of cell culture growth (398.94 ± 8.32 g/L Fresh cell weight (FCW) and 40.00 ± 0.75 g/L Dry cell weight (DCW) on 40-day as compared to control (MS with 3.0 mg/L 2, 4-d + 2.0 mg/L Kn). High-performance liquid chromatography analysis showed that maximum accumulation of deacylgymnemic acid (5.51 mg/g DCW), gymnemagenin (2.80 mg/g DCW) and gymnemic acid XVII (2.08 mg/g DCW) in 20 μM SNP treatment which is (13.43, 13.86 and17.33 folds) higher than the respective control at 40 days exposure. This research suggests that G. sylvestre cell suspension culture with optimal SNP elicitation treatment could be used as a good strategy for the large-scale production of these secondary metabolites at the industrial level.
... Similarly, decline biomass accumulation tendency was observed in Leucas aspera (Poornima et al., 2020). Cai et al. (2012), Salma et al. (2018), Zare-Hassani et al. (2019) and Poornima et al. (2020) had concluded that the effect of elicitors on growth of suspension culture depends on the type of elicitor, concentration, exposure time and investigation conditions. As previously described by Wang et al. (2015), Deepthi and Satheeshkumar (2016), Salma et al. (2018) and Udomsin et al. (2020) supplementation of elicitors during the log phase/ deceleration phase of suspension culture enhanced significantly and improved biomass production. ...
... Cai et al. (2012), Salma et al. (2018), Zare-Hassani et al. (2019) and Poornima et al. (2020) had concluded that the effect of elicitors on growth of suspension culture depends on the type of elicitor, concentration, exposure time and investigation conditions. As previously described by Wang et al. (2015), Deepthi and Satheeshkumar (2016), Salma et al. (2018) and Udomsin et al. (2020) supplementation of elicitors during the log phase/ deceleration phase of suspension culture enhanced significantly and improved biomass production. Our finding also showed that biomass enhancement was based on the concentration and incubation time of elicitor. ...
Gymnemic Acids (GAs) belong to the class of triterpenoid saponins present majorly in the leaves of Gymnema sylvestre. These bioactive compounds are responsible for hypoglycemic activity. Therefore, it is expansively used in Ayurveda treatment for diabetes. This study is aimed to develop an effective method for suspension culture and to investigate the effect of sodium nitroprusside (SNP), salicylic acid (SA) and methyl jasmonate (MeJA) on the enhancement of deacylgymnemic acid, gymnemagenin, gymnemic acid IV and gymnemic acid XVII in G. sylvestre cell suspension culture. To induce callus, in vitro-derived leaf explants were cultivated on Murashige and Skoog's (MS) medium supplemented with different concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D) or α-naphthalene-acetic acid (NAA) either alone or in combination with 6-benzylaminopurine (BAP) or kinetin (Kin) at 1.0 to 5.0 mg/L. The highest callus initiation frequency (95.30 ± 1.40%) and better friable callus observed on MS medium supplemented with 3.0 mg/L (2,4-D) + 1.0 mg/L (Kin). Optimized callus induction medium was used to develop cell suspension culture, growth kinetic study, elicitors treatment and quantify the production of major bioactive compounds. At 40 days, the highest fresh cell suspension culture biomass was 339.08 g/L fresh weight (FW). Biomass production, 366.68 g/L FW and 30.30 g/L DW were the highest in the treatment of 20 μM SNP among the tested elicitors. The highest accumulation of deacylgymnemic acid (2936.90 μg/g DW, 96 h treatment) and gymnemic acid XVII (2414.20 μg/g dry weight (DW), 48 h treatment) were obtained in 20 μM SNP treatments which were 6.1 and 5.2 fold higher than the control. Furthermore, the highest values of gymnemagenin (1179.77 μg/g DW at 50 μM treatment) and gymnemic acid IV (731.04 μg/g DW at 10 μM treatment) were obtained in SA treatment which was 4.12 and 5.11 fold greater than the control was studied through our newly developed HPLC method. The findings of the present study showed that the SNP could be a good strategy for large-scale industrial production of triterpenoid saponins in G. sylvestre cell suspension cultures.
... Earlier studies demonstrated that elicitors are very effective for the enhancement of cell growth and increase metabolites accumulation. Furthermore, the type of elicitors, concentration and exposure period favours the biomass accumulation in cell suspension culture (Salma et al., 2018;Zare-Hassani et al., 2019;Açıkgöz, 2020). ...
Gymnema sylvestre (Retz.) R.Br. ex Sm. is widely used as an efficient Ayurvedic traditional medicine for the treatment of diabetes. Phytochemical investigations of this plant showed gymnemic acids (a group of triterpenoid saponins) as the main active components. The present study aims to investigate the effectiveness of sodium nitroprusside (SNP) treatment for enhancement of cell suspension culture biomass and to evaluate their deacylgymnemic acid, gymnemagenin, gymnemic acid IV and gymnemic acid XVII contents of G. sylvestre . Callus was obtained from in vitro derived leaves of G. sylvestre on MS medium fortified with 3.0 mg/L 2, 4-D (2, 4-dichlorophenoxyacetic acid) and 2.0 mg/L Kn (Kinetin), and the same were used further to produce cell suspension cultures. Cell suspensions were exposed to different concentrations of SNP (5, 10, 20 and 40 µM) and data were collected at 20, 30 and 40 days. Out of the tested concentrations, 20 µM SNP had the highest level of cell culture growth (398.94 ± 8.32 g/L FCW and 40.00 ± 0.75 g/L DCW) on 40-day as compared to control. High-performance liquid chromatography analysis showed that maximum accumulation of deacylgymnemic acid (5.51 mg/g DCW), gymnemagenin (2.80 mg/g DCW) and gymnemic acid XVII (2.08 mg/g DCW) in 20 µM SNP treatment which is (13.43, 13.86 and17.33 folds) higher than the respective control at 40 days exposure. This research suggests that G. sylvestre cell suspension culture with optimal SNP elicitation treatment could be used as a good strategy for the large-scale production of these secondary metabolites at the industrial level.
... The findings obtained from this study showed that the plants exposed to IT contain higher concentration of wedelolactone as compared to CT and control. Elicitor-induced enhancement in wedelolactone concentration in cell suspension culture of E. alba were also reported by Salma et al. (2018). ...
In the present study sensitivity of a medicinal plant Eclipta alba L. (Hassk) (False daisy) was assessed under intermittent (IT) and continuous (CT) doses of elevated ultraviolet-B (eUV-B). Eclipta alba is rich in medicinally important phytochemical constituents, used against several diseases. The hypothesis of this study is that alterations in UV-B dose may modify the quantity and quality of medicinally valuable components with changes in the morphological and physiological parameters of test plant. To fulfill our hypothesis IT and CT of eUV-B (ambient ± 7.2 kJ m−2 day−2) was given for 130 and 240 h respectively to assess the impact of UV-B stress. Growth and physiological parameters were adversely affected under both the treatments with varying magnitude. The observation of leaf surfaces showed increase in stomatal and trichome densities suggesting the adaptive resilience of the plants against UV-B. Besides, biosynthesis of wedelolactone, a major medicinal compound of E. alba was observed to be stimulated under UV-B exposure. The essential oil content was reduced under IT while increased under CT. A total of 114 compounds were identified from oil extract of E. alba. n-Pentadecane (25.79%), n-Octadecane (12.98%), β-Farnesene (9.43%), α-Humulene (4.95%) (E)-Caryophyllene (4.87%), Phytol (4.25%), α-Copaene (2.26%), Humulene epoxide (1.46%), β-Pinene (1.07) and β-Caryophyllene oxide (1.06%) were identified as major components of oil. CT induced the synthesis of some medicinally important compounds such as α-terpineol, δ-cadinene, linolenic acid, methyl linoleate and myristic acid amide. Hence, the study revealed that continuous UV-B exposure of low intensity could be helpful for commercial exploitation of essential oil in E. alba.
... The setup of high-producing in vitro cultures is an important stage for obtaining the secondary metabolites from the plant cell cultures (Smetanska 2008). The addition of biotic and abiotic elicitors to the cells culture elicits production of secondary metabolite in plant cells and it considers as an enhancement strategy (Simic et al. 2015;Ramirez-Estrada et al. 2016;Salma et al. 2018). Even plant cells engineered to overexpress biosynthetic pathway key genes yet need elicitation for high production of related secondary metabolite. ...
Paclitaxel is an impressive chemotherapeutic agent that shows biological activity against a wide range of cancers. Cell suspension culture of Corylus avellana has been reported as an alternative source for production of the valuable secondary metabolite, paclitaxel. The fungal elicitors have been considered as the most impressive strategy for enhancement of secondary metabolites in plant cell culture. In this study, two endophytic fungal strains YEF20 and HEF114 were isolated from Taxus baccata and C. avellana, respectively. The isolates YEF20 and HEF114 were identified as Chaetomium globosum and Paraconiothyrium brasiliense, respectively by sequencing of ITS1-5.8S-ITS2 rDNA region. This is the first report of P. brasiliense on C. avellana tree. Also, this study presents the positive effect of fungal elicitors on paclitaxel production in C. avellana for the first time. The effect of fungal elicitors on paclitaxel production was dependent on fungal species, and also type, concentration and adding time of elicitors to cell culture. The highest total yield of paclitaxel in cell culture treated with cell extract (CE) of C. globosum (291.5 µg L⁻¹) was obtained by using 10% (v/v) of this fungal elicitor on 17th day of cell culture cycle (late log-phase), which was about 4.1 times obtained in the control culture. The cell culture treated with C. globosum CE had an average growth rate of 0.491 g L⁻¹ day⁻¹, i.e. 12.3% lower than that in the control. Among the fungal elicitors, 10% (v/v) CE of C. globosum on 17th day of culture cycle showed the best results with respect to extracellular paclitaxel portion (44.0%). Paclitaxel secretion to culture medium is essential for large-scale steady production of paclitaxel.
Plants are one of the most reliable resources of nutrient supplements and pharmaceutical compounds, but rapidly losing from the wild due to intensified harvesting for the bioactive secondary metabolites of nutraceutical interest. Recent resurgence in the nutraceutical and its importance in the human health has aroused worldwide researchers to find efficient techniques for nutraceutical production. Among others, in vitro plant cell culture technique is a viable alternative for large-scale propagation of germplasm with the high content of nutraceutical compounds. However, for successful in vitro plant cell culture, selection of a suitable plant material and optimization of several physical and chemical culture conditions such as nutrient media content, mineral composition, phytohormone supplementation, temperature, pH, light, humidity, aeration, agitation, etc. is imperative. The optimization of these parameters accelerates the growth rate of plant cells under in vitro conditions along with the improvement of production of nutraceuticals. Therefore, the present chapter provides details on the optimization of in vitro plant cell culture methods with respect to sustainable production of nutraceutical compounds.
The present investigation aimed to improve callus biomass, polyphenolic content, biosynthesis of mangiferin and biological potential following application of different elicitor treatments for medicinally important Salacia chinensis L. The leaf-derived callus cultures were established on Murashige and Skoog’s (MS) medium supplemented with 2,4-dichlorophenoxyacetic acid (2,4-D: 2.0 mg/l) and 6-benzylaminopurine (BAP: 1.5 mg/l). These cultures were treated with different elicitors viz. jasmonic acid (JA), methyl jasmonate (MeJA) and yeast extracts (YE). The highest calli biomass (five-fold increase within 4 weeks) was achieved when callus was treated with JA (75 µM). The callus obtained on MS medium supplemented with 2,4-D (2.0 mg/l), BAP (1.5 mg/l) and treated with JA (75 µM) displayed augmented values for total phenolics, flavonoids and mangiferin contents. Besides, same treatment elicits the calli for antioxidant properties as evaluated by 2,2-diphenyl-2-picrylhydrazyl (DPPH), ferric-reducing antioxidant power (FRAP) and metal chelating assays. This is the first report on the elicitation study in genus Salacia and, therefore, the discoveries suggested that, S. chinensis calli might be a perfect source for large-scale production of industrially important secondary metabolites. Concurrently data provide accumulated information demonstrating its prominent antioxidant effect revealing its potential without disturbing natural resources.
